Assay for p53 function in cells
First Claim
1. A method for identifying compounds that modulate the binding of p53 protein to a gene of interest, comprising:
- providing said gene of interest as chromatin-assembled DNA;
contacting said gene of interest with said p53 protein and a test compound; and
determining whether the presence of said test compound modulates the binding of said p53 to said gene of interest.
1 Assignment
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Accused Products
Abstract
Disclosed herein is an assay for identifying compounds that modulate the interaction between p53 and specific DNA binding sites. The assay uses chromatin-assembled target genes to correctly reproduce p53 binding to target genes of interest. This assay permits the selection of compounds which preferentially associate with p53 when bound to specific DNA binding sites but not others. Moreover, compounds can be screened for their ability to selectively modulate p53 interaction with specific binding sites associated with individual genes. Compounds can also be screened for their ability to selectively modulate p53 interaction with specific cofactors (proteins, nucleic acids, peptides, etc.) when p53 is bound to specific DNA sequences. This system allows drugs to be tailored to affect p53-dependent regulation of certain target genes that regulate distinct cellular pathways without affecting other genes and pathways.
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Citations
35 Claims
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1. A method for identifying compounds that modulate the binding of p53 protein to a gene of interest, comprising:
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providing said gene of interest as chromatin-assembled DNA;
contacting said gene of interest with said p53 protein and a test compound; and
determining whether the presence of said test compound modulates the binding of said p53 to said gene of interest. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12)
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13. A method for identifying a modulator that inhibits growth of cancer cells, comprising:
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providing a gene construct comprising a chromatin-assembled promoter linked to a reporter gene, wherein said promoter binds p53 protein;
incubating said gene construct in the presence of p53 protein and a test compound; and
determining whether said test compound increases binding of said p53 protein to said gene construct by measuring the amount of reporter gene activity in the presence and the absence of said test compound. - View Dependent Claims (14, 15, 16, 17, 18)
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19. A method for identifying a test compound that modulates the interaction of p53 protein with a transcription cofactor that associates with p53 protein when p53 protein is bound to a gene of interest, comprising:
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providing said gene of interest as chromatin-assembled DNA;
contacting said gene of interest with said p53 protein, the transcription cofactor, and the test compound; and
determining whether the presence of said test compound modulates the association of said cofactor with said p53 protein bound to said gene. - View Dependent Claims (20, 21, 22, 23, 24, 25, 26, 27)
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28. A kit for identifying test compounds that affect p53 protein binding to a promoter, comprising:
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a gene construct comprising a chromatin assembled p53 binding site genetically linked to a reporter gene; and
isolated p53 protein. - View Dependent Claims (29, 30, 31, 32, 33, 34, 35)
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Specification